To investigate the functional role of different α₁-adrenergic receptor (α₁-AR) subtypes in vivo, we have applied a gene targeting approach to create a mouse model lacking the α_1b-AR (α_1b−/−). Reverse transcription–PCR and ligand binding studies were combined to elucidate the expression of the α₁-AR subtypes in various tissues of α_1b +/+ and −/− mice. Total α₁-AR sites were decreased by 98% in liver, 74% in heart, and 42% in cerebral cortex of the α_1b −/− as compared with +/+ mice. Because of the large decrease of α₁-AR in the heart and the loss of the α_1b-AR mRNA in the aorta of the α_1b−/− mice, the in vivo blood pressure and in vitro aorta contractile responses to α₁-agonists were investigated in α_1b +/+ and −/− mice. Our findings provide strong evidence that the α_1b-AR is a mediator of the blood pressure and the aorta contractile responses induced by α₁ agonists. This was demonstrated by the finding that the mean arterial blood pressure response to phenylephrine was decreased by 45% in α_1b −/− as compared with +/+ mice. In addition, phenylephrine-induced contractions of aortic rings also were decreased by 25% in α_1b−/− mice. The α_1b-AR knockout mouse model provides a potentially useful tool to elucidate the functional specificity of different α₁-AR subtypes, to better understand the effects of adrenergic drugs, and to investigate the multiple mechanisms involved in the control of blood pressure.